title: Molecular characterization of a glycosyl hydrolase family 10 xylanase from Aspergillus niger.
abstract: A gene coding for an endo-β-1,4-xylanase (XlnA) (glycosyl hydrolase family 10) from Aspergillus niger DSM 1957 was cloned and sequenced. The cDNA sequence (984 bp) and its putative endoxylanase (327 aa protein with a predicted molecular mass of 35.5 kDa and pI 6.23) showed 91.3-99.5% and 96.3-99.1% identities with cDNA sequences and their corresponding endoxylanases from A. niger strains from GenBank, respectively. The cDNA was expressed in Pichia pastoris GS115 under the control of AOX1 promoter at a level of 46.4 U/ml culture supernatant, after 144 h of growth at 30°C in YP medium induced with 0.5% (v/v) of methanol. The molecular mass of the purified XlnA determined by SDS-PAGE was 35.5k Da with a specific activity of 808.5 U/mg towards 1% (w/v) of birch wood xylan. Temperature and pH optimum were observed at 50°C and pH 7.0, respectively. The enzyme was stable over a temperature range of 25-40°C and at pH range of 4.5-8.5 and resistant to Tween 80 and acetone. The K(m) and V(max) value obtained for the purified xylanase were 25.5mg/ml and 5000 μmol/min/mg protein with birch wood xylan as substrate, respectively. The xylanase was free of cellulase and mannanase activity but highly active towards birch wood xylan. The major products of the birch wood xylan hydrolysis were predicted as xylotriose, xylotetraose, and xylopentose. The biochemical characteristics suggested that the recombinant xylanase has a potential application, including use as a feed enzyme.
authors.affiliation: Institute of Biotechnology, Viet Nam Academy of Science and Technology, 18 Hoang Quoc Viet Road, Cau Giay Distr., 10600 Hanoi, Viet Nam.
authors.forename: Thi Tuyen
authors.initials: TT
authors.lastname: Do
authors.forename: Dinh Thi
authors.initials: DT
authors.lastname: Quyen
authors.forename: Thi Nuong
authors.initials: TN
authors.lastname: Nguyen
authors.forename: Van Thuat
authors.initials: VT
authors.lastname: Nguyen
mesh_terms: D001234:Aspergillus niger / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D003902:Detergents / Q000494:pharmacology; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008670:Metals / Q000494:pharmacology; D011994:Recombinant Proteins / Q000737:chemistry* / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D012997:Solvents / Q000494:pharmacology; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D003902:Detergents; D008670:Metals; D011994:Recombinant Proteins; D012997:Solvents; D043364:Endo-1,4-beta Xylanases
keywords: Aspergillus niger DSM 1957; Cloning; Expression; Glycosyl hydrolase family 10 xylanase; Properties